The present invention relates to a polymer-coated capillary tube used in an electrophoresis apparatus, to a method for processing a window at a predetermined position of the capillary tube for enhancing a light transmittance thereof and to a capillary tube whose coating is selectively removed to provide a window by the method.
Electrophoresis apparatuses that utilize capillary tubes for rapid processing of mass data are widely used in DNA nucleotide sequence analyses. The capillary tubes used are generally glass tubes having an inner diameter of 1 to 800 xcexcm, an outer diameter of 50 to 1,000 xcexcm and a length of 50 to 2,000 mm, which is coated with a polymer to a thickness of 10 to 30 xcexcm, among which a capillary tube of an appropriate size is employed. Polymer, generally polyimide, is coated to enhance the mechanical strength and give flexibility to the fragile glass tube for easy handling. However, since a sample migrating inside a capillary tube is detected by irradiating the fluorescence-labeled sample with a laser beam from outside the capillary tube and detecting the excited fluorescence, a polymer coating with a poor light transmittance will interfere with the sample detection. Thus, it is necessary to selectively remove the polymer coating for a length of about 1 to 20 mm around the periphery of the capillary tube. The portion removed of the polymer coating for an increased light transmittance is generally called a xe2x80x9cwindowxe2x80x9d.
In order to process a window for a capillary tube, a technique in which a polymer coating is removed by burning with a lighter flame is generally employed when the number of capillary tubes is small. When the number of the capillary tubes is large, a technique such as one disclosed in JP-A-5-232085 may be employed in which a plurality of capillary tubes are supported at the same time to burn and remove predetermined regions of the polymer coating with a gas burner. However, burning with a lighter or a gas burner will leave cinders on the surfaces of the capillary tubes, and thus requires an after-treatment for wiping the cinders away with a wiper such as paper or cloth, which causes an increase in the production cost. Furthermore, wiping with the wiper may damage the exposed glass tube region. If the exposed glass tube region has a damage, the capillary tube can be susceptible to breakage during operations such as mounting the capillary tube on an apparatus. In addition, controlling the temperature of the flame is difficult since burning at a low temperature will leave a large amount of cinders and burning at a high temperature will result in deformation or poor transmittance of the glass capillary tube. Cinders on the window will interfere with the detection of fluorescence excited by laser irradiation.
In order to process a window by burning the polymer coating under a controlled temperature, techniques using an electric heater as disclosed in U.S. Pat. No. 4,940,883 (1990), JP-A-10-206383, JP-A-11-230939 may be employed. However, even under a controlled temperature, polymer coating is difficult to be burnt completely at or below an allowable continuous temperature of synthesized quartz (about 950xc2x0 C.) that is generally used for making the glass tube, and a border between regions heated and unheated by the electric heater always includes insufficiently heated parts. As a result, after-treatment for wiping away the cinders is inevitable. Even if the window is elegantly processed, remaining cinders may fall off after the capillary tube is mounted on the electrophoresis apparatus, and may stick to a part that may cause an adverse effect on detection.
As a processing technique other than burning, JP-A-6-74938 discloses a technique for mechanically removing the coating with a scraper. However, it is difficult to remove the coating efficiently without damaging the glass tube. According to another technique, the polymer coating, will be sublimed through ablation with an excimer laser. However, this technique requires an expensive device and a large installation area. Compounds such as hot sulfuric acid and hydrazine may be used for removing the coating. However, they are hazardous and thus have drawbacks such as difficult handling, requirement of after-treatment, requirement of waste fluid processing and a risk of causing adverse effects on the environment.
A capillary tube provided with a window is susceptible to breakage because stresses caused upon bending or pulling the capillary tube may concentrate on the damage of the exposed glass region. The polymer coating is provided basically to enhance the mechanical strength and flexibility of the glass tube. Another reason for easy breakage of the capillary tube by the concentration of the stresses on the exposed portion is the profile of the edge of the coating at the window. When the edge of the coating at the window is removed generally perpendicular to the longitudinal direction of the capillary tube, the stresses will be concentrated on the edge of the coating at the window upon bending the tube and cause easy breakage.
In a multi-capillary electrophoresis apparatus, a plurality of capillary tubes are used as a multi-capillary array with the windows arranged in parallel to form a plane with the ends of the tubes being aligned. The windows of the multi-capillary array are supported by a holder so that the tubes are held secure. Capillary tubes having windows are used to produce the multi-capillary array. Since the mechanical strength of the glass capillary tubes with exposed windows is poor, they are difficult to be handled upon assembly, which interferes with automation of producing a multi-capillary array.
The above-described conventional techniques have adverse effects on the strength and the measurement due to damage, deformation, poor transmittance or the like of a glass capillary tubes and have the following problems: requirement of an after-treatment following the coating removal; requirement of an expensive device and a large installation area for the device; or use of a compound that may be hazardous to humans and the environment. In addition, conventional techniques have a problem of being susceptible to breakage which is caused by concentration of stresses on the edge of the coating at the window of the capillary tube, and a problem of automation in assembling fragile capillary tubes having windows being impossible for producing a multi-capillary array.
The present invention has an objective of providing a tough capillary tube provided with a window, which is produced in a safe and inexpensive manner by a method that causes no adverse effect, such as damage, deformation, poor transmittance or the like on a glass tube as a main body of the capillary tube, which may interfere with optical detection of a sample by an electrophoresis apparatus and while saving trouble such as wiping after the coating removal. The present invention also has an objective of providing means for automating production of a multi-capillary array, which has been impossible.
The above-mentioned objectives are achieved by applying a reactive gas such as ozone to a selective region of a capillary tube where a window is to be processed. Specifically, according to the present invention, one or more capillary tubes are placed in a reaction chamber such that a desirable window-processing length of about 1 to 20 mm of the capillary tubes is exposed inside the space of the reaction chamber. Then, a reactive gas containing ozone is supplied to the space inside the reaction chamber. The parts of the capillary tubes in the space as well as the reactive gas are heated so that the ozone contained in the reactive gas is decomposed to generate oxygen radical. Only the polymer coating on the parts exposed to the space inside the reaction chamber will be converted into gas and removed through an oxidative reaction with the oxygen radical.
According to this method, windows can be processed by removing the polymer coating at a much lower temperature (400xc2x0 C. or lower) than a heat deformation temperature of the capillary tubes. By heating as well as radiating ultraviolet ray to the capillary tubes and the reactive gas filling the space inside the reaction chamber, ozone will absorb the ultraviolet ray (254 nm) to be more susceptible to decomposition. Thus, the windows can be processed through polymer coating removal at a lower temperature (250xc2x0 C. or lower). In order to remove only the coatings on the predetermined regions of the capillary tubes, the plurality of capillary tubes are arranged perpendicular to the flow of the reactive gas while only the window-processing parts are surrounded by the semi-sealed reactive unit. Since the capillary tubes are supported by a holder, windows can be processed without the risk of damaging the tubes.
As shown in FIGS. 3 to 6, the reaction chamber is a hollowed space formed between and at generally the center of two substrates sandwiching the capillary tubes. One of the two substrates is provided with one or more grooves for supporting the capillary tubes. The reaction chamber is provided with a supply port and a discharge port for the reactive gas, which are provided at opposite sides of and appropriately apart from the space of the reaction chamber. The reactive gas introduced from the supply port into the space of the reaction chamber flows across the surfaces of the capillary tubes in the reaction chamber and discharged from the discharge port.
The capillary tubes are heated through heat transfer by mounting the two substrates sandwiching the capillary tubes on a heating plate above the device body. The temperature of the capillary tubes is lower than the maximum heat resistant temperature of polyimide (material of the polymer coating). When the reactive gas makes contact with the heated capillary tubes, the temperature of the capillary tubes is decreased and the oxidative reaction is suppressed. Therefore, the reactive gas is pre-heated to achieve a temperature generally equal to the temperature of the capillary tubes immediately before the reactive gas makes contact with the capillary tubes, thereby shortening the time required for processing the windows. The reactive gas is pre-heated by providing a heater or the like at the pipe immediately in front of the reactive gas supply port, by extending the distance between the supply port and the capillary tubes so that the reactive gas is heated to a temperature generally equal to that of the capillary tubes by the heating unit for the capillary tubes, or by transferring the heat from the heating plate from the reactive gas supply port to a pipe extending to an ozone generator so that the reactive gas is pre-heated to a temperature generally equal to that of the capillary tubes while flowing through the pipe.
Removal of the polymer coating using the reactive gas containing ozone takes place by supplying ozone to the heated polymer coating, by which the ozone will make contact with the heated capillary tubes or the ozone will be pre-heated by the pre-heating mechanism to be decomposed into O2 and O radical, thereby promoting the oxidative reaction. However, the coating is not removed when either heating or reactive gas supply is lacking. In order not to remove the polymer coating on the capillary tubes other than the parts in the reaction chamber, dimensional tolerances of the width and the depth of the grooves for supporting the capillary tubes are slightly larger than the size of the capillary tubes to allow a slight amount of gas to flow between the capillary tubes and the grooves supporting the capillary tubes. As a result, a slight amount of atmosphere will be drawn in the space inside the reaction chamber and the ozone will make contact with the capillary tubes only into the space inside the reaction chamber. This structure allows the length of the processed windows to be generally equal to the length of the parts of the capillary tubes exposed to the space inside the reaction chamber. Accordingly, the length of the windows to be processed can be adjusted by varying the size of the reaction chamber. Since a slight amount of atmosphere is drawn into the reaction chamber from the gap between the capillary tubes and the grooves, the ozone is supplied to the space inside the reaction chamber not by a pressure higher than the atmospheric pressure being applied to the supply port but by being drawn into the space through negative pressure by making the pressure at the discharge port lower than atmospheric pressure. This structure also serves as a seal to prevent the ozone from leaking outside the device.
By making the upper substrate of the two substrates sandwiching the capillary tubes from glass with a high light transmittance, the removal status of the polymer coating on the capillary tubes can be observed. Therefore, the termination of the window-processing operations can be determined or the capillary tubes can be heated with a lamp heater or a laser beam through the glass substrate. By radiating ultraviolet ray to the capillary tubes and the reactive gas filling inside the reaction chamber, the ozone will absorb the ultraviolet ray (254 nm) to be susceptible to decomposition, which allows removal of the polymer coating (window-processing) at a lower temperature (250xc2x0 C. or lower).
The present invention applies the above-mentioned structures to produce a multi-capillary array in a simple manner. Specifically, the holder (made of non-polymer material in this case) for covering and holding parts of the multi-capillary array to prevent them from falling apart is used as a reaction chamber for processing the windows. According to this method, first, a multi-capillary array is assembled, and then they are provided with windows. Therefore, the multi-capillary array can be produced by using capillary tubes having no window, which are stronger and easier to handle, thereby greatly enhancing an operation efficiency. Furthermore, production of the multi-capillary array can be automated.
In order to remove the polymer coating on the capillary tubes, other than the reaction chamber, an ozone generator, an ozone killer for decomposing the remaining unreacted ozone, a power source, a controller and the like are necessary. These members can be connected via pipes or lines to be used in a window-processing device for capillary tubes. According to the present invention, each of the constituent members is optimized and accommodated in a single body to produce a device that is specialized in processing windows for capillary tubes, which can be used on a desk.